Symbiotic interactions among microorganisms are abundant in nature. The unusual combination of genetic, biochemical and chemical techniques required to study these interactions has hampered their detailed analysis, and therefore most remain poorly-examined. One of the most abundant and environmentally important symbioses occurs in the oceans between microscopic alga, like Emiliania huxleyi, and bacteria of the roseobacter clade, such as Phaeobacter gallaeciensis. E. huxleyi occupies all sun-lit ocean layers and plays an important role in global oxygen and carbon cycles. It forms massive seasonal blooms, where it intermittently associates with members of the roseobacter clade. Roseobacter are ubiquitous in coastal areas and play a major role in global sulfur cycles. While roseobacter-algal symbioses drive numerous biogeochemical processes, the molecular principles underlying these interactions remain unknown. Our preliminary results have shown that P. gallaeciensis, depending on circumstances, produces a potent, novel metabolite that kills E. huxleyi. The proposed research plan aims to 1) discover global regulators and small molecule signals that mediate or modulate roseobacter-algal interactions, 2) use NMR-based methods to characterize the structures of secondary metabolites produced by roseobacter in response to algal signals, and use bioassays to determine their functions, 3) delineate the biosynthetic pathway of these metabolites by transposon mutagenesis, gene deletions, and enzymatic studies, and 4) uncover how metabolite production is regulated using a combination of genetic and biochemical approaches. Subsequently, these studies will be extended to other roseobacter to examine the generality of the principles uncovered with E. huxleyi and P. gallaeciensis. This research plan will generate the tools needed to characterize many similar environmentally important interactions. Because symbioses contain a poorly-explored reservoir of metabolites with potential pharmaceutical and/or agricultural applications, this proposal could also identify novel and useful molecules. Harvard Medical School offers an intellectual niche and an established research program in this area or work. It consists of leaders in the fields of natural products chemistry and bacterial genetics who will serve as my mentors in the proposed project. Having obtained my PhD in mechanistic enzymology, my short-term goals are to acquire the skills necessary to examine the various aspects of microbial symbioses. In the mentored phase, I will be trained in bacterial genetics, small molecule characterization and relevant bioassays. During this time, I will also attend an advanced bacterial genetics course and other workshops/conferences to learn the scientific techniques and management skills required to be a successful PI. In the independent phase, these methods will be used to uncover the regulation of metabolite production and to examine the biosynthetic enzymes. In the long-term, I plan to lead a multidisciplinary research program in an academic institution to study the underlying chemistry, enzymology and biology of environmentally important symbioses. PUBLIC HEALTH RELEVANCE: A majority of today's pharmaceutical drugs were isolated from natural sources, such as plants and bacteria. To address the need to discover new molecules with improved properties, we will explore the compounds produced by ocean bacteria in response to the algae they interact with, and we will also address the chemical and biological mechanisms through which this interaction occurs. The studies will reveal the general principles governing these unexplored bacterial-algal symbioses and will hopefully yield novel compounds with pharmaceutical value.